| After the partial thickness burn, the involved skin always shows postburn hypopigmentation or hyperpigmentation, this phenomenon is often found in exposing sites such as face and hand, thus has heavy influences to the look of patients, while there is no ideal treatment so far duing to the poor understanding of its mechanism.Recently the transplantation of melanocytes cultured in vitro was used as a good measure to treat the hypopigmentation. However, because melanocytes constitute a minor component of the cell population in normal epidermis, and undergo very little replication, the in vitro cultures of melanocytes were always frustrated by the overgrowth of keratinocytes or fibroblasts. Traditionally some additives were used to suppress the unwanted cells, e.g. 12-o-tetradecanoyl-phorbol-13-acetate(TPA) was added to suppress the growth of keratinocytes, and geneticin to suppress the fibroblasts. But these additives have some by-effects, particularly, TPA is a tumor-promoting agent, the transplantation of melanocytes cultured by TPA has the possibility of tumor-promoting, with the investigation of paracrine regulation of melanocytes, basic fibroblast growth factor (bFGF) and some other factors have been found to be natural mitogens of melanocytes, thus can be applied to the in vitro culture. Because bFGF is a nature factors, the transplantation of melanocytes cultured by bFGF to treat the postburn hypopigmentation will be more safer than traditional measure.By far there are few reports about the treating of postburn hyperpigmentation, the effects of current treatments are not satisfactory. Arbutin has been found to be effective to some hyperpigmenting lesion, such as and jwithout by-effect, but there is no report about its effect in the treating of postburn hyperpigmentation. On the other hand, duing to the various cell strains and culture measures involved, there were significant discrepancies in some reports about the effects and mechanism of arbutin.The culture using bFGF as main mitogen is more similar to the in vivo environment of normal melanocytes, thus the results will be more believable if this system is used to study the depigmenting effects of arbutin.In the present study, bFGF and cholera toxin (CT) were used to obtain a pure culture of melanocytes, then the biologic characteristics of cultured.melanocytes were observed, including the morphology and proliferation, as well as the optimal parameters in subculturing, reserving and anabiosis. We also observed the effect of melanocytes transplantation in treating postburn hypopigmentation. Using this culture system, we studied the depigmenting effects of Arbutin, the cell viability was investigated by MTT assay, the tyrosinase activity was assessed utilizing L-Dopa as the substrate, the melanogenesis was assayed by the absorption in 490nm. Then we observed the effects of 5% Arbutin on postburn hyperpigmentation. The results were as follows:1.A pure culture of normal human epidermal melanocytes was obtained with a medium added bFGF and CT, the contamination of keratinocytes and fibroblasts was obviated. The optimal density in primary culture is 2xl05/cm2, while in subculture is IxlOVcnr, melanocytes can be repassaged 7-8 times, cells senesced about 7 weeks later, the total cell number doubled about 11 times, the mean population doubling time was about 5 days. The cultured melanocytes can resist liquid nitrogen abundantly with 60% cells being viable after anabiosis.2. Arbutin reduced cell viability about 7% at Img/ml, while caused no change in cell viability when lower than O.lmg/ml; arbutin showed a concentration-dependent reduction in tyrosinase activity at noncytotoxic concentrations, 50% inhibition of cellular tyrosinase activity was observed at Img/ml, while caused no change in tyrosinase activity when lower than O.OOlmg/ml; the melanogenesis of melanocytes was inhibited 14% by arbutin at 0.0 Img/ml.3. Kojic acid inhibited the cell viability 37% and 23% at Img/ml and O.lmg/ml , respectively, while caused no changes in cell viability...
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